Antibacterial composition

ABSTRACT

The present invention relates to an antibacterial composition and to a method for the treatment of infectious diseases caused by pathogenic bacteria employing a phosphonic acid derivative and an antibiotic.

CROSS REFERENCE TO RELATED APPLICATION

This is a continuation-in-part of copending application Ser. No. 877,944filed Feb. 15, 1978.

DETAILED DESCRIPTION

The present invention relates to a new antibacterial composition and toa new method for the treatment of infectious diseases caused bypathogenic bacteria. More particularly, it relates to an antibacterialcomposition comprising a phosphonic acid derivative of the formula:##STR1## wherein R is lower alkanoyl and

n is an integer of 2 to 5

or its salt and an antibiotic selected from a β-lactam antibiotic,aminoglycoside antibiotic and their salts,

and to a method for the treatment of infectious disease caused bypathogenic bacteria by application of said antibacterial composition toinfected human being or other animals.

As a result of extensive study of the present inventors, it has beennewly found that the phosphonic acid derivative (I) or its salt exhibitsa synergistic antibacterial activity by combination with an antibioticselected from a β-lactam antibiotic such as 1,3-disubstitutedazetidinone (e.g. nocardicin A), penicillin compound and cephalosporincompound, and aminoglycoside antibiotic (e.g. gentamicin), that is, thecombination of the phosphonic acid derivative (I) or its salt with anantibiotic selected from the β-lactam antibiotic, aminoglycosideantibiotic and their salts shows an effectively stronger antimicrobialactivity against pathogenic bacteria in human being and other animals,against which the phosphonic acid derivative (I), the β-lactamantibiotic, or aminoglycoside antibiotic shows no or less antibacterialactivity enough to effectively treat human being and other animals forinfectious diseases or to effectively prevent them from said diseases,when each of said compounds is used alone.

Accordingly, an object of the present invention is to provide anantibacterial composition comprising the phosphonic acid derivative (I)or its salt and an antibiotic selected from a β-lactam antibiotic,aminoglycoside antibiotic and their salts, which is useful as anantimicrobial agent against pathogenic bacteria in human being and otheranimals, against which each of the phosphonic acid derivative (I), theβ-lactam antibiotic, the aminoglycoside antibiotic and their salts aloneis not so effective.

Another object of this invention is to provide a method for thetreatment of infectious diseases caused by pathogenic bacteria, whichcomprises administering the phosphonic acid derivative (I) or its saltin combination with an antibiotic selected from a β-lactam antibiotic,aminoglycoside antibiotic and their salts.

These and other objects of the present invention will be apparent fromthe description hereinafter.

The antibacterial composition of the present invention comprises acombination of the phosphonic acid derivative (I) or its salt and anantibiotic selected from a β-lactam antibiotic, an aminoglycosideantibiotic and their salts.

With regard to the phosphonic acid derivative (I) to be used in thisinvention, preferred "lower alkanoyl" for R is one having 1 to 6 carbonatoms, among which the most preferred one is formyl and acetyl; and aninteger of 3 is most preferred for the symbol "n". That is, thephosphonic acid derivative of the following formula (I') is the mostpreferred compound to be used in this invention. ##STR2## wherein R' isformyl and acetyl.

The phosphonic acid derivative (I) is an antibiotic having antibacterialactivity against various pathogenic bacteria and can be produced byfermentation and/or synthesis, the details of which are described inBelgian Pat. No. 857.211, in which the preferred phosphonic acidderivative (I') can be prepared, for example, by culturing Streptomycesrubellomurinus ATCC 31215 or Streptomyces lavendulae ATCC 31279 in anutrient medium and recovering the produced3-(N-acetyl-N-hydroxyamino)propylphosphonic acid or3-(N-formyl-N-hydroxyamino)propylphosphonic acid from the resultantcultured broth, respectively.

Further, the β-lactam antibiotic to be used in this invention includes1,3-disubstituted azetidinone, penicillin and cephalosporin compounds,and the preferred β-lactam antibiotic is nocardicin A for1,3-disubstituted azetidinone, ampicillin, carbenicillin and ticarcillinfor penicillin, and cefazolin for cephalosporin, which are famousantibiotics described in e.g. THE JOURNAL OF ANTIBIOTICS Vol. 29, pages492-500 (1976), THE MERCK INDEX NINTH EDITION pages 80 and 228 (1976),ANTIMICROBIAL AGENTS AND CHEMOTHERAPY Vol. 7, pages 336-340 (1975), andTHE MERCK INDEX NINTH EDITION pages 245 (1976), respectively.

Furthermore, the aminoglycoside antibiotic to be used in this inventionincludes gentamicin, which is a famous antibiotic described in e.g. THEMERCK INDEX NINTH EDITION pages 565-566 (1976).

The salts of the above antibiotics, e.g. the phosphonic acid derivative(I), β-lactam antibiotic such as nocardicin A, ampicillin,carbenicillin, ticarcillin and cefazolin, and aminoglycoside antibioticsuch as gentamicin may include physiologically (e.g. pharmaceutically)acceptable salts such as a metal salt (e.g. sodium, potassium, calcium,barium or magnesium salt), ammonium salt, an amine salt (e.g.ethanolamine, triethylamine, procaine, dibenzylamine ordicyclohexylamine salt), an acid addition salt (e.g. sulfate) and thelike.

From the above description, it is to be noted that the preferredcombination of the phosphonic acid derivative (I) and an antibioticselected from the β-lactam antibiotic and aminoglycoside antibiotic is acombination of the phosphonic acid derivative (I') and an antibioticselected from nocardicin A, ampicillin, carbenicillin, ticarcillin,cefazolin and gentamicin.

The antibacterial composition of the present invention is useful fortreating and preventing infectious diseases induced by pathogenicbacteria in human being and other animals such as poultry, domesticanimals, pet animals or experimental animals (e.g. chicken, turkey,duck, quail, cow, cattle, horse, pig, hog, dog, sheep, goat, mink,canary, macaw, mouse, rat or rabbit).

The combination ratio of the phosphonic acid derivative (I) or its saltand an antibiotic selected from the β-lactam antibiotic, theaminoglycoside antibiotic and their salts in the present antibacterialcomposition may vary depending on the kinds of pathogen and the symptomsof the patients to which the present composition is applied, but mayusually be selected within a range of 1:4 to 4:1 by weight, preferably1:2 to 2:1 by weight and most preferably 1:1.

Further, it is to be noted that the present antibacterial compositionmay be applied to human being and other animals in conventional forms,examples of which are illustrated as follows.

For applying the present antibacterial composition to human, it ispreferable to apply it in the form of intravenous or intramuscularinjection. It may also be applied locally in the form of a powder, asuppository or an ointment. When used as an injection, it may be appliedin admixture with a solid or liquid carrier or diluent which is usuallyused for the conventional antibiotic injections, and further, may alsobe applied together with other medicines such as analgesics (e.g.lidocaine) which are usually used in injections. The most preferredcarrier or diluent is water. When used as a suppository and an ointment,it may be used in admixture with conventional suppository and ointmentbases, respectively.

For applying the present antibacterial composition to other animals, itis preferable to apply it in the form of injection or in the form ofinfusion. It may also be applied locally in a form of a powder or anointment. When used as an injection or infusion, it may be applied inadmixture with a solid or liquid carrier or diluent which is usuallyused for the conventional antibiotic injections or infusions. The mostpreferred carrier or diluent is water, vegitable oils, paraffins or thelike. When used as an ointment, it may be applied in admixture withconventional ointment bases.

The dosage of the present antibacterial composition may vary dependingon the kinds of the phosphonic acid derivative (I), the β-lactamantibiotic and aminoglycoside antibiotic, the combination ratio thereofand various factors such as the weight and age of the patient, the kindand severity of the infection, and the kind of the application mode.However, it is to be understood that, as the dosage of the effectiveingredient included in the present antibacterial composition, it may beeffectively administered to the patient in a dose of about 5 to 200mg/kg/day, preferably 10-100 mg/kg/day in the case of a combination ofthe phosphonic acid derivative (I) and the β-lactam antibiotic and in adose of about 0.3-5 mg/kg/day in the case of a combination of thephosphonic acid derivative (I) and β-lactam antibiotic. Moreparticularly, for instance, in the injection with a combination of thephosphonic acid derivative (I) and β-lactam antibiotic to human being,it may be administered in a dose of about 1 to 5 g/day in adults and ina dose of about 10 to 30 mg/kg/day in children, but not limited thereto.For treating bovine mastitis during lactation drying period, acombination of the phosphonic acid (I) and the β-lactam antibiotic maybe administered in a dose of about 50-500 mg/quarter. The total dailyamount mentioned above may be divisionally given to the patient at theinterval of 6-12 hours per day.

And further, it is to be noted that the present antibacterialcomposition shows low toxicity as shown in the following toxicity test.

ACUTE TOXICITY TEST

The acute toxicity test was conducted by using each of the followingantibacterial composition according to the following experimentalprocedure.

(1) Antibacterial composition

(a) Composition of monosodium salt of3-(N-acetyl-N-hydroxyamino)propylphosphonic acid and monosodium salt ofnocardicin A (1:1 by weight).

(b) Composition of monosodium salt of3-(N-formyl-N-hydroxyamino)propylphosphonic acid and monosodium salt ofnocardicin A (1:1 by weight).

(c) Composition of monosodium salt of3-(N-formyl-N-hydroxyamino)propylphosphonic acid and monosodium salt ofampicillin (1:1 by weight).

(d) Composition of monosodium salt of3-(N-formyl-N-hydroxyamino)propylphosphonic acid and monosodium salt ofcefazolin (1:1 by weight).

(e) Composition of monosodium salt of3-(N-formyl-N-hydroxyamino)propylphosphonic acid and monosodium salt ofticarcillin (1:1 by weight).

(f) Compositions of monosodium salt of3-(N-formyl-N-hydroxyamino)propylphosphonic acid and disodium salt ofcarbenicillin (1:1 by weight)

(2) Experimental procedure

An aqueous solution (0.5 ml) containing one of the above antibacterialcomposition was intravenously injected into each of three ICR-strainmale mice weighing 20 g (Dose: 500 mg/kg), respectively. The observationwas continued for one week after the administration.

(3) Test results

All of the test mice were living and normal.

Further, as to a composition of monosodium salt of3-(N-formyl-N-hydroxyamino)propylphosphonic acid and gentamicin sulfate(1:1 by weight), an acute toxicity test was conducted in substantiallythe same manner as described above, excepting Dose: 5 mg/kg of mouse.The test result obtained was the same as the above Test results.

The antibacterial activities and the preventing effectiveness againstvarious bacterial infections of the present antibacterial compositionare illustrated in the following experimental tests in vitro and invivo.

Test 1

Synergistic activity of 3-(N-acetyl-N-hydroxyamino)propylphosphonic acidand nocardicin A in vitro:

Into a Nutrient broth (Difco) containing prescribed amount of each ofmonosodium salt of 3-(N-acetyl-N-hydroxyamino)propylphosphonic acid,monosodium salt of nocardicin A and a mixture of monosodium salt of3-(N-acetyl-N-hydroxyamino)-propylphosphonic acid and monosodium salt ofnocardicin A (1:1 by weight), there was inoculated overnight-culturedbroth of each pathogen in a final concentration of 10⁶ cells/ml,respectively. After the incubation was carried out at 37° C. for 20hours, Minimum Inhibitory Concentration (MIC) values were determined,respectively.

Further, in order to observe the degree of synergistic antimicrobialactivity, Fractional Inhibitory Concentration (FIC) values and FIC Indexwere calculated from the determined MIC values according to thefollowing calculation method, respectively.

Calculation method

(a) MIC value of monosodium salt of3-(N-acetyl-N-hydroxyamino)propylphosphonic acid: Ao

(b) MIC value of monosodium salt of nocardicin A: Bo

(c) MIC value of a mixture of monosodium salt of3-(N-acetyl-N-hydroxyamino)propylphosphonic acid and monosodium salt ofnocardicin A: Cab

On the basis of the fact that the combination ratio of a mixture ofmonosodium salt of 3-(N-acetyl-N-hydroxyamino)propylphosphonic acid andmonosodium salt of nocardicin A is 1:1 (by weight), each of FIC valuesand FIC indexes was calculated according to the following equations.

FIC of monosodium salt of 3-(N-acetyl-N-hydroxyamino)propylphosphonicacid=(1/2 Cab/Ao).

FIC of monosodium salt of nocardicin A=(1/2 Cab/Bo).

FIC index=(1/2 Cab/Ao)+(1/2 Cab/Bo).

The test results are shown in the following table.

                  Table 1.                                                        ______________________________________                                        Synergism between 3-(N-acetyl-N-hydroxyamino)propylphos-                      phonic acid and nocardicin A against pathogenic bacteria                      Micro-  strain  MIC (mcg/ml)  FIC      FIC                                    organism                                                                              No.     A       B    C    A     B    index                            ______________________________________                                        Pseudo- 1101-63 800     400  100  0.063 0.125                                                                              0.188                            monas   1101-64 >800    200  50   <0.031                                                                              0.125                                                                              <0.156                           aeruginosa                                                                            1101-66 200     100  50   0.125 0.250                                                                              0.375                                    1101-67 200     200  100  0.250 0.250                                                                              0.500                                    1101-68 800     100  50   0.031 0.250                                                                              0.281                            Escherichia                                                                           1341-27 200     100  50   0.125 0.250                                                                              0.375                            coli    1341-35 100     100  50   0.250 0.250                                                                              0.500                            Klebsiella                                                                            1391-1  400     800  100  0.125 0.063                                                                              0.188                            pneumoniae                                                                            1391-2  >800    800  400  <0.250                                                                              0.250                                                                              <0.500                                   1391-3  800     800  400  0.250 0.250                                                                              0.500                                    1391-5  400     800  200  0.250 0.125                                                                              0.375                            Serratia                                                                              1421-1  >800    100  12.5 <0.008                                                                              0.063                                                                              <0.071                           marcescens                                                                            1421-3  >800    400  100  <0.063                                                                              0.125                                                                              <0.188                                   1421-4  >800    400  50   <0.031                                                                              0.063                                                                              <0.094                                   1421-5  >800    200  25   <0.016                                                                              0.063                                                                              < 0.079                          ______________________________________                                         (Note)                                                                        A: Monosodium salt of 3(N-acetyl-N-hydroxyamino)propylphosphonic acid.        B: Monosodium salt of nocardicin A.                                           C: A mixture of monosodium salt of                                            3(N-acetyl-N-hydroxyamino)propylphosphonic acid and monosodium salt of        nocardicin A (1:1 by weight).                                            

Test 2

Synergistic activity of 3-(N-formyl-N-hydroxyamino)propylphosphonic acidand ampicillin in vitro:

On a Nutrient agar (Difco) containing prescribed amount of each ofmonosodium salt of 3-(N-formyl-N-hydroxyamino)propylphosphonic acid,monosodium salt of ampicillin and a mixture of3-(N-formyl-N-hydroxyamino)propylphosphonic acid and monosodium salt ofampicillin (1:1 by weight), there was streaked a loopful of culturedbroth of each pathogen, which was cultured overnight in Nutrient broth(Difco), in a concentration of 10⁵ cells/ml. After the incubation wascarried out at 37° C. for 20 hours, MIC values were determined.

FIC values and FIC Index were calculated in substantially the samemanner as described in Test 1.

The test results are shown in the following table.

                  Table 2.                                                        ______________________________________                                        Synergism between 3-(N-formyl-N-hydroxyamino)propylphos-                      phonic acid and ampicillin against pathogenic bacteria                                         MIC (mcg/ml)                                                                              FIC                                              Microorganism      A       B      C    index                                  ______________________________________                                        Staphylococcus epidermidis 1601-1                                                                ≧400                                                                           0.2    0.2  0.5                                    Klebsiella pneumoniae NCTC 418                                                                   25      12.5   3.13 0.2                                    Shigella flexneri Ia EW-8                                                                        1.56    0.78   0.78 0.8                                    Salmonella enteritidis 1891                                                                      0.78    0.39   0.39 0.8                                    Salmonella typhimurium 1406                                                                      6.25    0.39   0.39 0.5                                    Salmonella paratyphi A-1015                                                                      6.25    0.39   0.39 0.5                                    Serratia marcescens 1421-4                                                                       50      25     3.13 0.1                                    Enterobacter aerogenes 1402-10                                                                   6.25    200    3.13 0.3                                    Enterobacter clocae 1401-4                                                                       3.13    100    3.13 0.5                                    Alcaligenes faecalis 1311-1                                                                      ≧400                                                                           6.25   6.25 0.5                                    Proteus mirabilis 1432-75                                                                        1.56    0.39   0.2  0.3                                    Proteus vulgaris IAM-1025                                                                        1.56    0.78   0.78 0.8                                    Proteus morganii 1433-2                                                                          ≧400                                                                           25     25   0.5                                    ______________________________________                                         (Note)                                                                        A: Monosodium salt of 3(N-formyl-N-hydroxyamino)propylphosphonic acid.        B: Monosodium salt of ampicillin                                              C: A mixture of monosodium salt of                                            3(N-formyl-N-hydroxyamino)propylphosphonic acid and monosodium salt of        ampicillin (1:1 by weight).                                              

Test 3

Synergistic activity of 3-(N-formyl-N-hydroxyamino)propylphosphonic acidand cefazolin; that of 3-(N-formyl-N-hydroxyamino)propylphosphonic acidand ticarcillin; and that of 3-(N-formyl-N-hydroxyamino)propylphosphonicacid and nocardicin A were tested in vitro in substantially the samemanner as described in the above Test 2.

The test results are shown in the following Table 3, 4 and 5,respectively.

                  Table 3.                                                        ______________________________________                                        Synergism between 3-(N-formyl-N-hydroxyamino)propylphos-                      phonic acid and cefazolin against pathogenic bacteria                                          MIC (mcg/ml) FIC                                             Microorganism      A       B       C    index                                 ______________________________________                                        Staphylococcus epidermides 1601-1                                                                ≧400                                                                           0.39    0.39 0.5                                   Escherichia coli 1341-18R.sup.+                                                                  6.25    3.13    1.56 0.4                                   Klebsiella pneumoniae NCTC 418                                                                   25      1.56    1.56 0.5                                   Serratia marcescens 1421-4                                                                       50      400     25   0.3                                   Enterobacter aerogenes 1402-10                                                                   6.25    12.5    3.13 0.4                                   Enterobacter cloacae 1401-4                                                                      3.13    ≧400                                                                           3.13 0.5                                   Proteus rettgeri 1434-3                                                                          1.56    6.25    1.56 0.6                                   Proteus morganii 1433-2                                                                          ≧400                                                                           25      25   0.5                                   Pseudomonas aeruginosa 1101-76                                                                   1.56    ≧400                                                                           1.56 0.5                                   ______________________________________                                         (Note)                                                                        A: Monosodium salt of 3(N-formyl-N-hydroxyamino)propylphosphonic acid.        B: Monosodium salt of cefazolin.                                              C: A mixture of monosodium salt of                                            3(N-formyl-N-hydroxyamino)propylphosphonic acid and monosodium salt of        cefazolin (1:1 by weight).                                               

                  Table 4.                                                        ______________________________________                                        Synergism between 3-(N-formyl-N-hydroxyamino)propylphos-                      phonic acid and ticarcillin against pathogenic bacteria                                        MIC (mcg/ml)                                                                              FIC                                              Microorganism      A       B      C    index                                  ______________________________________                                        Staphylococcus epidermides 1601-1                                                                ≧400                                                                           3.13   3.13 0.5                                    Klebsiella pneumoniae NCTC418                                                                    25      100    25   0.6                                    Salmonella typhimurium 1406                                                                      6.25    1.56   1.56 0.6                                    Serratia marcescens 1421-4                                                                       50      25     12.5 0.4                                    Enterobacter cloacae 1401-4                                                                      3.13    50     3.13 0.5                                    Proteus mirabilis 1432-75                                                                        1.56    0.78   0.78 0.8                                    ______________________________________                                         (Note)                                                                        A: Monosodium salt of 3(N-formyl-N-hydroxyamino)propylphosphonic acid         B: Monosodium salt of ticarcillin                                             C: A mixture of monosodium salt of                                            3(N-formyl-N-hydroxyamino)propylphosphonic acid and monosodium salt of        ticarcillin (1:1 by weight)                                              

                  Table 5.                                                        ______________________________________                                        Synergism between 3-(N-formyl-N-hydroxyamino)propylphos-                      phonic acid and nocardicin A against pathogenic bacteria                                       MIC (mcg/ml) FIC                                             Microorganism      A      B      C      index                                 ______________________________________                                        Klebsiella pneumoniae NCTC418                                                                    25     200    25     0.6                                   Salmonella typhimurium 1406                                                                      6.25   25     6.25   0.6                                   Serratia marcescens 1421-4                                                                       50     25     6.25   0.2                                   Proteus mirabilis 1432-75                                                                        1.56   3.13   0.78   0.4                                   ______________________________________                                         (Note)                                                                        A: Monosodiuim salt of 3(N-formyl-N-hydroxyamino)propylphosphonic acid        B: Monosodium salt of nocardicin A                                            C: A mixture of monosodium salt of                                            3(N-formyl-N-hydroxyamino)propylphosphonic acid and monosodium salt of        nocardicin A (1:1 by weight)                                             

Test 4

Synergistic activity of 3-(N-formyl-N-hydroxyamino)propylphosphonic acidand carbenicillin in vitro:

Onto a Nutrient agar (Difco) containing prescribed amount of each ofmonosodium salt of 3-(N-formyl-N-hydroxyamino)propylphosphonic acid,disodium salt of carbenicillin, and a mixture of3-(N-formyl-N-hydroxyamino)propylphosphonic acid and disodium salt ofcarbenicillin (1:1 by weight), there was spot-inoculated, using amultiple inoculator cultured broth of each pathogen, which was culturedovernight in Nutrient broth (Difco), in a concentration of 10⁸ cells/ml.After the incubation was carried out at 37° C. for 20 hours, MIC valueswere determined.

FIC values and FIC Index were calculated in substantially the samemanner as described in Test 1.

The test results are shown in the following table.

                  Table 6.                                                        ______________________________________                                        Synergism between 3-(N-formyl-N-hydroxyamino)propylphos-                      phonic acid and carbenicillin against pathogenic bacteria                                     MIC (mcg/ml) FIC                                              Microorganism     A       B       C    index                                  ______________________________________                                        Pseudomonas aeruginosa No.5                                                                     800     50      6.25 0.067                                  Pseudomonas aeruginosa No.7                                                                     200     >800    25   <0.078                                 Pseudomonas aeruginosa No.9                                                                     >800    >800    100  <0.125                                 Pseudomonas aeruginosa No.14                                                                    400     >800    50   <0.094                                 Pseudomonas aeruginosa No.15                                                                    >800    800     50   <0.063                                 Escherichia coli No.14                                                                          25      12.5    6.25 0.38                                   ______________________________________                                         (Note) A: Monosodium salt of 3(N-formyl-N-hydroxyamino)propylphosphonic       acid.                                                                         B: disodium salt of carbenicillin                                             C: A mixture of monosodium salt of                                            3(N-formyl-N-hydroxyamino)propylphosphonic acid and disodium salt of          carbenicillin (1:1 by weight).                                           

Test 5

Synergistic activity of 3-(N-formyl-N-hydroxyamino)propylphosphonic acidand gentamicin in vitro:

Onto a Nutrient agar (Difco) containing prescribed amount of each ofmonosodium salt of 3-(N-formyl-N-hydroxyamino)propylphosphonic acid,gentamicin sulfate and a mixture of3-(N-formyl-N-hydroxyamino)propylphosphonic acid and gentamicin sulfate(1:1 by weight), there was spot-inoculated, using a multiple inoculatorcultured broth of each pathogen, which was cultured overnight inNutrient broth (Difco), in a concentration of 10⁸ cells/ml. After theincubation was carried out at 37° C. for 20 hours, MIC values weredetermined.

FIC values and FIC Index were calculated in substantially the samemanner as described in Test 1.

The test results are shown in the following table.

                  Table 7.                                                        ______________________________________                                        Synergism between 3-(N-formyl-N-hydroxyamino)propylphos-                      phonic acid and gentamicin against pathogenic bacteria                                        MIC (mcg/ml)                                                                              FIC                                               Microorganism     A       B      C    index                                   ______________________________________                                        Pseudomonas aeruginosa No.5                                                                     800     50     25   0.27                                    Pseudomonas aeruginosa No.11                                                                    >800    3.13   1.56 <0.25                                   Pseudomonas aeruginosa No.12                                                                    800     12.5   3.13 0.13                                    Pseudomonas aeruginosa No.13                                                                    800     100    50   0.28                                    Pseudomonas aeruginosa No.18                                                                    >800    50     25   <0.27                                   ______________________________________                                         (Note)                                                                        A: Monosodium salt of 3(N-formyl-N-hydroxyamino)propylphosphonic acid         B: Gentamicin sulfate                                                         C: A mixture of monosodium salt of                                            3(N-formyl-N-hydroxyamino)propylphosphonic acid and gentamicin sulfate        (1:1 by weight)                                                          

As seen clearly from the above results in Test 1-5, the combination ofthe phosphonic acid derivative(I) and an antibiotic selected fromNocardicin A, ampicillin, ticarcillin, carbenicillin, cefazolin andgentamicin shows synergistic antibacterial activity against variouspathogens.

Test 6

Effects on the experimentally infected mice:

ICR-strain male mice weighing 23-25 g (10 mice per one group) were used.A prescribed amount of the pathogenic bacteria suspended in 5% aqueousmucin suspension (0.5 ml.) was inoculated intraperitoneally into eachmouse. One hour after the inoculation, the antibiotics as mentioned inthe following table were administered subcutaneously, and then survivalof the test mice was measured one week after the infection to determineED₅₀ value. The results are shown in the following Table 8.

Incidentally, FIC values and FIC index in this in vivo test were alsocalculated from the determined ED₅₀ values according to the followingcalculation method.

Calculation method

(a) ED₅₀ value of monosodium salt of3-(N-acetyl-N-hydroxyamino)propylphosphonic acid: A'o

(b) ED₅₀ value of monosodium salt of nocardicin A:B'o

(c) ED₅₀ value of a mixture of monosodium salt of3-(N-acetyl-N-hydroxyamino)propylphosphonic acid and monosodium salt ofnocardicin A:C'ab

On the basis of the fact that the combination ratio of a mixture ofmonosodium salt of 3-(N-acetyl-N-hydroxyamino)propylphosphonic acid andmonosodium salt of nocardicin A is 1:1 (by weight), each of FIC valuesand FIC index was calculated according to the following equations.

FIC of monosodium salt of 3-(N-acetyl-N-hydroxyamino)propylphosphonicacid=(1/2C'ab)/A'o.

FIC of monosodium salt of nocardicin A=(1/2C'ab)/B'o.

FIC index=(1/2C'ab)/A'o+(1/2C'ab)/B'o.

                  Table 8.                                                        ______________________________________                                        Synergism between 3-(N-acetyl-N-hydroxyamino)propylphos-                      phonic acid and nocardicin A in protecting activity                           Micro-   strain  ED.sub.50 (mg/mouse)                                                                       FIC      FIC                                    organism No.     A      B    C    A     B    index                            ______________________________________                                        Pseudomonas                                                                            1101-5  >20    1.3  0.71 <0.018                                                                              0.273                                                                              <0.291                           aeruginosa                                                                             1101-7  >20    15.0 2.5  <0.063                                                                              0.083                                                                              <0.146                           ______________________________________                                         (Note) A: Monosodium salt of 3(N-acetyl-N-hydroxyamino)propylphosphonic       acid                                                                          B: Monosodium salt of nocardicin A                                            C: A mixture of monosodium salt of                                            3(N-acetyl-N-hydroxyamino)propylphosphonic acid and monosodium salt of        nocardicin A (1:1 by weight)                                             

As seen clearly from the above results, the synergistic antibacterialactivity of 3-(N-acetyl-N-hydroxyamino)propylphosphonic acid andnocardicin A was also confirmed by in vivo test.

The antibacterial compositions of the present invention are illustratedby the following Examples.

EXAMPLE 1

A sterile mixture of monosodium salt of3-(N-formyl-N-hydroxyamino)propylphosphonic acid (125 mg.) andmonosodium salt of nocardicin A (125 mg.) was put in a sterile vial andthe vial was sealed. And when used, the above mixture was dissolved in asterile water (2 ml.) to give an injection preparation.

In substantially the same manner as described in the above Example 1,there was prepared an injection preparation of an antimicrobialcomposition as illustrated in the following Examples 2-7.

EXAMPLE 2

A mixture of monoammonium salt of3-(N-acetyl-N-hydroxyamino)propylphosphonic acid (250 mg.) andmonosodium salt of nocardicin A (125 mg.) was used as the activeingredient for injection.

EXAMPLE 3

A mixture of monopotassium salt of3-(N-formyl-N-hydroxyamino)propylphosphonic acid (250 mg.) andmonosodium salt of ampicillin (250 mg.) was used as the activeingredient for injection.

EXAMPLE 4

A mixture of monosodium salt of3-(N-formyl-N-hydroxyamino)propylphosphonic acid (125 mg.) andmonosodium salt of cefazolin (125 mg.) was used as the active ingredientfor injection.

EXAMPLE 5

A mixture of monosodium salt of3-(N-formyl-N-hydroxyamino)propylphosphonic acid (250 mg.) andmonosodium salt of ticarcillin (250 mg.) was used as the activeingredient for injection.

EXAMPLE 6

A mixture of monosodium salt of3-(N-formyl-N-hydroxyamino)propylphosphonic acid (250 mg.) and disodiumsalt of carbenicillin (250 mg.) was used as the active ingredient forinjection.

EXAMPLE 7

A mixture of monosodium salt of3-(N-formyl-N-hydroxyamino)propylphosphonic acid (20 mg) and gentamicinsulfate (20 mg) was used as the active ingredient for injection.

What is claimed is:
 1. An antibacterial composition comprising (a) aphosphonic acid derivative of the formula: ##STR3## wherein R is formylor acetyl, or its pharmaceutically acceptable salt and (b) an antibioticselected from ampicillin, carbenicillin, ticarcillin or theirpharmaceutically acceptable salts, wherein (a) and (b) are contained inthe composition in a ratio of 1:2 to 2:1 by weight.
 2. The antibacterialcomposition of claim 1 which comprises (a) 3-(N-formyl-N-hydroxyamino)propylphosphonic acid or its pharmaceutically acceptable salt and (b)ampicillin or its pharmaceutically acceptable salt.
 3. The antibacterialcomposition of claim 1 which comprises (a) 3-(N-formyl-N-hydroxyamino)propylphosphonic acid or its pharmaceutically acceptable salt and (b)ticarcillin or its pharmaceutically acceptable salt.
 4. A method oftreating an infectious disease caused by a pathogenic bacteria in humansand animals, which comprises administering thereto an effective amountof the antibacterial composition of claim
 1. 5. A method of treating aninfectious disease caused by a pathogenic bacteria in humans andanimals, which comprises intravenously or intramuscularly administratingthereto a daily dose of 10 to 100 mg/kg, as effective ingredient of theantibacterial composition of claim
 1. 6. A method of treating aninfectious disease caused by a pathogenic bacteria in humans andanimals, which comprises intravenously or intramuscularly administratingthereto a daily dose of 0.3 to 5 mg/kg, as effective ingredient of theantibacterial composition of claim
 1. 7. An antibacterial compositioncomprising 3-(N-formyl-N-hydroxyamino)propylphosphonic acid monosodiumsalt, and carbenicillin disodium salt in a ratio 1 to 1 by weight.
 8. Amethod of treating an infectious disease caused by a pathogenic bacteriain humans and animals, which comprises intravenously or intramuscularlyadministrating thereto a daily dose of 10 to 100 mg/kg, as effectiveingredient of the antibacterial composition of claim
 7. 9. Anantibacterial composition comprising 3-(N-formyl-N-hydroxyamino)propylphosphonic acid monosodium salt, and carbenicillin disodium saltin a ratio 1:2 to 2:1 by weight.